Pulmonary surfactant is critically important for normal lung function. Surfactant deficiency is the cause of the Respiratory Distress Syndrome of the Newborn (RDS), a condition which has been estimated to contribute to 20% of all neonatal deaths. Alterations in this material also contribute to forms of Adult Respiratory Distress (ARDS) that are consequent to metabolic or chemical insult to the lung. However, it is not known which of the steps in the overall synthesis or processing of surfactant are affected in the various pathological conditions that result in respiratory distress. The alveolar type II cell is the source of disaturated phosphatidylcholine, the major component of surfactant. This proposal will utilized cell line A549, which has been demonstrated by our previous work to be a cell culture model of the type II pneumocyte. We will define the major stages in the production of surfactant phospholipid in A549 cells in order to determine the points at which physiological and pathological effectors might act to cause an alteration. Two complementary and interrelated approaches will be used: 1. We will examine selected aspects of phospholipid biochemistry in A549 cells and subcellular fractions which pertain to surfactant production. Specifically, we will examine qualitative and quantitative lipid composition, and precursor incorporation pathways, kinetics and enzymatic activities involved in phosphatidylcholine synthesis. 2. Since cellular secretory processes are highly vectorial in nature, it is essential to understand the ultrastructural aspects of surfactant phospholipid synthesis and translocation. We have previously shown that an organometallic analog of palmitate (a tin-containing fatty acid) is incorporated into surfactant-like phospholipid synthesized by A549 cells and that this prove can be detected by electron microscopy and X-ray microanalysis. The proposed studies will further develop this labeling strategy and apply it to the examination of the ultrastructural dynamics of disaturated phosphatidylcholine synthesis.